Background:Opioids are the principal drugs of choice for managing acute severe pain; however, physical dependence is still reported as one of the main limiting factors in the clinical application of these drugs. In the present study, the effect of Ganoderma lucidum (G. lucidum) was assessed on morphine dependence in mice.
Methods:A 19-day administration schedule was applied to induce morphine dependence in male adult NMRI mice. The mice were given intraperitoneal (i.p.) morphine sulfate once daily in an increasing dose of 10, 20, and 40 mg/kg. Then, G. lucidum hydroalcoholic extract (12.5, 25, and 50 mg/kg, i.p.) was given to the mice from days 10 to 18. Another group of mice received single doses of the extract (50, 100, and 200 mg/kg, i.p.) only on the 19th day. Naloxone (3 mg/kg, i.p.) was used to precipitate withdrawal syndrome. Normal saline and diazepam (0.25 mg/kg) were used as the negative and positive controls, respectively.
Results:The administration of single doses of G. lucidum extract (100 and 200 mg/kg, i.p.) significantly decreased the number of jumps, leanings, and diarrhea in mice subjected to morphine dependence. The repeated administration of G. lucidum extract (25 and 50 mg/kg for nine days) significantly attenuated the number of jumps, leanings, and diarrhea in morphine-dependent mice.
Conclusions:Overall, G. lucidum extract attenuates induced morphine dependence and inhibits withdrawal syndrome symptoms in mice.
Physical dependence and addiction are still the main dilemmas in the clinical application of opioid analgesic drugs (1). Many endeavors have been made to resolve the problem of opioid dependence by focusing on its underlying physiopathological mechanisms (2-5). In this regard, it has been demonstrated that benzodiazepine/GABA receptors (6), calcium currents (7, 8), oxidative/nitrosative stress (9, 10), and glial cell-activated neuroinflammation (11) play an essential role in the pathophysiology of opioid dependence. Accordingly, several investigations have indicated that applying naturally occurring compounds, including herbal medicines, might be worthwhile for managing dependence on opioid agents by targeting the aforementioned mechanisms (12-15).
Ganoderma lucidum (G. lucidum), a member of the Ganodermataceae family, is a significant source of several natural medicinal mushrooms with a long history of therapeutic applications for the management of many diseases (16-18). Ganoderma lucidum has suitable therapeutical value for the isolation of new bioactive compounds. Triterpenoids and polysaccharides are the primary bioactive metabolites isolated from G. lucidum and have a vital role in its pharmacological effects (19). A 12.4 kDa immunomodulatory protein (Ling Zhi-8) have described from G. lucidum mycelia with biological activity (20). Ganoderma lucidum has also shown anti-tumor, antimicrobial, and anti-aging activities (21-23).
Several pharmacological effects have been known for G. lucidum. For instance, evidence demonstrates that G. lucidum exhibits sedative and sleep-promoting properties (24-26) through activating benzodiazepine/GABAA receptors (27). Furthermore, polysaccharides extracted from G. lucidum represented an inhibitory effect on intracellular calcium accumulation in hippocampal neurons (28). Moreover, G. lucidum extract and its major constituents, including polysaccharides and triterpenes, have an inhibitory effect against oxidative/nitrosative stress (29-31). Besides, recent studies demonstrated the inhibitory effect of G. lucidum on glial cells and inflammatory cytokines (31-33).
Considering the pharmacological effects of G. lucidum on benzodiazepines/GABAA receptors, calcium currents, oxidative/nitrosative stress, and neuroinflammation and the fact that these mechanisms contribute to the induction of opioid dependence, it was hypothesized that G. lucidum might attenuate morphine dependence by targeting the aforementioned mechanisms. Thus, the present study aimed to clarify the effects of G. lucidum hydroalcoholic extract on the induction of morphine dependence and the expression of morphine abstinence syndrome in mice.
Adult male NMRI mice with a weight of 20 to 30 g, which were purchased from the animal house of Shahid Beheshti University of Medical Sciences (Tehran, Iran), were used in this study. The mice were housed under standard conditions at a temperature of 23±2°C and a 12 h light/dark cycle. The animals were fed commercial food pellets and had free access to tap water, except during the experiments. All the procedures performed on the animals, including handling, restraining, and injecting, were carried out under the institutional guidelines for laboratory animal care and use.
3.2. Process of Drug Preparation
3.2.1. Extraction Method
The percolation route was chosen to produce a hydroalcoholic extract of G. lucidum. Hence, 50.2 g of the fine powder of the biomass was moved into a glassy percolator, 5 cm in diameter and 30 cm in height. Then, 150 mL of methanol/water (7:3) solvent was transferred to the percolator. After 24 h, its valve was opened, and the filtered extract was carried out at one droplet per second. This process was repeated another three times
3.2.2. Condensation and Dehumidification of the Extract
The extract was condensed using a rotary evaporator (Heidolph, Germany) at 40°C. The final condensed extract was dried with a lab heater (Heidolph, Germany) at 40°C. The dried extract was stored at 4°C for subsequent steps.
Morphine sulfate and naloxone hydrochloride were purchased from Temad Co. (Iran) and Tolidaru Co. (Iran), respectively. Normal saline was used as the vehicle to solve morphine and naloxone. All the injection solutions were prepared freshly and administered through the intraperitoneal (i.p.) route (injection volume: 0.1 mL/ 20 g body weight). In the negative control group, the same volume of normal saline was administered i.p. in mice. Diazepam (Caspian Co., Iran) was used as the positive control.
3.4. Assessment of G. lucidum Effect on Morphine Dependence
A 19-day administration schedule was implemented to induce morphine dependence. First, the mice were given morphine sulfate, i.p., once daily over nine days in an increasing dose of 10, 20, and 40 mg/kg so that each dose was administered for three days. Then, the i.p. injection of morphine at 40 mg/kg was continued over the next 10 days, i.e., days 10 to 19 (34).
To evaluate the effect of G. lucidum on the development of morphine dependence, we divided the mice into four groups (six in each group), including one control group and three G. lucidum-treated groups. In the treatment groups, the mice received different doses of G. lucidum hydroalcoholic extract (12.5, 25, and 50 mg/kg, i.p.), while in the control group, normal saline was injected into the animals from days 10 to 18.
To evaluate the effect of G. lucidum on the expression of morphine dependence, we divided the mice into four groups (six in each group), including a control group and three G. lucidum-treated groups. In the treatment groups, three single doses of G. lucidum (50, 100, and 200 mg/kg, i.p.) were administered to the mice, while in the control group, normal saline was given to the animals on the 19th day.
One hour after the injection of the last dose of morphine on the 19th day, all mice received a single dose of naloxone (3 mg/kg, i.p.) and were immediately placed in a Plexiglass box. Then, the symptoms of morphine abstinence syndrome, including jumping, leaning, and diarrhea, were assessed in each mouse for 30 min (14).
3.5. Statistical Analysis
The data were reported as mean values ± SEM for six mice per group and analyzed using one-way ANOVA, followed by the multiple comparison test of Tukey. If the P-value was less than 0.05, the result was considered significant.
4.1. Yield of Extraction
When a solid material comes in contact with a solvent, the soluble components in the solid material move into the solvent (35); thus, the final extract had a 4.51 g weight, and the efficiency of the extracting process was 8.98%.
4.2. Effects of G. lucidum on Morphine Dependence
The administration of multiple doses of morphine over 19 days resulted in a remarkable dependence in the control group's mice. This outcome was confirmed by the symptoms of morphine withdrawal syndrome, which was precipitated by injection of naloxone (3 mg/kg, i.p.) one hour after the last dose of morphine on the 19th day.
Treatment of mice with single doses of G. lucidum extract (100 and 200 mg/kg, i.p.) one hour before the injection of naloxone on the 19th day significantly decreased the number of jumps (P < 0.01 and P < 0.001, respectively) and leanings (P < 0.05 and P < 0.001, respectively). The high dose of G. lucidum extracts significantly decreased diarrhea in mice subjected to morphine dependence (P < 0.01) (Figure 1).
The repeated administration of G. lucidum extracts with doses of 25 and 50 mg/kg, i.p., for nine days significantly attenuated the number of jumps (P < 0.05 and P < 0.001, respectively) and leanings (P < 0.05) in morphine-dependent mice (Figure 2).
As the positive control, administering single or repeated diazepam doses (0.25 mg/kg, i.p.) significantly suppressed the symptoms of morphine withdrawal syndrome, including jumps, leaning, and diarrhea in morphine-dependent mice (Figures 1 and 2).
The present study demonstrated for the first time that G. lucidum extracts inhibited the induction of morphine dependence and suppressed the morphine abstinence syndrome in mice. The attenuating effect of G. lucidum on withdrawal syndrome was as effective as the standard medication, diazepam, which was applied as the positive control.
In the present study, the chronic administration of increasing doses of morphine for 19 days induced apparent physical dependence in mice which was evident by the symptoms of withdrawal syndrome elicited by naloxone injection. This protocol is one of the most fundamental ways of inducing morphine dependence in mice and rats by chronically administering morphine (34, 36). Although the acute administration of morphine in a three-day injection schedule is commonly applied to induce morphine dependence (14, 15), the long administration pattern further resembles the pathophysiological process of morphine dependence. In the present study, the treatment of mice with G. lucidum for 10 days significantly attenuated the symptoms of morphine withdrawal syndrome. Furthermore, the injection of single doses of G. lucidum on day 19 considerably decreased the naloxone-precipitated morphine withdrawal syndrome. The effect of both single and multiple doses of G. lucidum on morphine dependence in mice was dose-dependent and comparable to diazepam.
Evidence indicates that G. lucidum exhibits sedative and sleep-promoting properties (24-26, 37, 38). These studies suggested that the sedative-hypnotic effects of G. lucidum might be attributed to an agonistic effect on benzodiazepine/GABAA receptors (27). It has been demonstrated that GABAergic neurons are essential in modifying opioid dependence. For instance, benzodiazepines as the positive modulators of benzodiazepine receptors and the GABAA receptor agonist, muscimol, can prevent morphine dependence and attenuate the signs and symptoms of deprivation syndrome in morphine-dependent rodents (6, 39-41). Considering the involvement of benzodiazepines and GABAA receptors in the therapeutic effects of G. lucidum, it appears that the attenuating effects of G. lucidum on morphine dependence and the prevention of morphine withdrawal syndrome are associated with its stimulatory effect on benzodiazepine/GABAA receptors. Our result is consistent with previous studies indicating that GABA-modulating medicinal herbs efficiently attenuate morphine dependence in rodents (42-44).
Furthermore, the contribution of calcium currents and the role of intracellular calcium has been confirmed in opioid dependence (7, 8, 45). Evidence indicates that polysaccharides in G. lucidum have an inhibitory effect on the accumulation of intracellular calcium in epileptic hippocampal neurons (28). Thus, it can be argued that the attenuating effect of G. lucidum on morphine dependence is exerted at least in part through its inhibitory effect against calcium accumulation inside the neurons.
Ample evidence shows that G. lucidum extract and its major constituents, particularly polysaccharides and triterpenes, possess a potent inhibitory effect against oxidative/nitrosative stress through enhancing the antioxidant enzyme activities, preventing lipid peroxidation, scavenging free radicals, and inhibiting nitric oxide (NO) production (29-31, 46-48). Evidence indicates that ethanol extract of G. lucidum increases cellular antioxidant potential via activation of Nrf2 (49). Phytochemical analysis has shown ganoderic acids A, B, C, and D, ganodermanontriol, lucidenic acid B, and fatty acid amides (oleamide, hexadecanamide, and 9-oxo-10 Octadecadienoic acid) are major chemicals that function as antioxidant agents in G. lucidum extracts (47). Several studies demonstrated that oxidative and nitrosative stress is vital in the induction and development of opioid dependence (9, 10, 50). Align with these reports, it has been demonstrated that antioxidant medications, including medicinal herbs, have a promising effect in treating opioid dependence (51, 52). Accordingly, it seems that the suppressive effect of G. lucidum on morphine dependence partly refers back to its inhibitory effects on oxidative/nitrosative stress.
Recently, Torkzadeh-Mahani et al. described the contribution of morphine to glial cell-activated neuroinflammation in the CNS, which may result in morphine dependence (53). Also, Zhang et al. argued that activated astrocytes and microglial cells could promote mechanisms that underlie opioid dependence and addiction (54). There is evidence indicating that inflammatory cytokines secreted from astrocytes and microglial cells induce the process of central sensitization and consequently reduce the therapeutic effects of morphine (55). The inflammatory cytokines seem essential in acquiring physical dependence on morphine and morphine deprivation syndrome (11). Based on the importance of glial cell-activated neuroinflammation in eliciting opioid dependence, it is conceivable to accept that the inhibition of neuroinflammation in the CNS through suppressing activated glial cells and inhibiting inflammatory cytokines is a feasible strategy for attenuating opioid dependence. In this regard, several studies demonstrated the anti-inflammatory effect of G. lucidum (33) and confirmed that it has an inhibitory effect on astrocytes and microglial cells, as well as inflammatory cytokines (31, 32, 56). Ethnopharmacological studies showed that triterpenoids and steroids, including ganoderic acid C and 3-oxo-5α-lanosta-8,24-dien-21-oic acid, are the main components of G. lucidum with anti-inflammatory properties (57, 58) (Joseph et al., 2009; Ko et al., 2008). Thus, it appears that G. lucidum can attenuate morphine dependence, at least in part, by inhibiting glial cells and inflammatory cytokines in the CNS.
Recently, it has been reported that G. lucidum extracts attenuated memory impairment induced by morphine and diminished conditioned place preference score in morphine-addicted mice (59). The present study, as many research works discussed earlier, deals with the therapeutic aspect of G. lucidum on morphine dependence by applying behavioral experiments in mice. However, the molecular mechanisms of G. lucidum action on morphine dependence should be further elucidated in future studies.
Overall, it is concluded that G. lucidum extract attenuates the induction of morphine dependence and inhibits the withdrawal syndrome symptoms in mice.
Alldredge BK, Corelli RL, Ernst ME, Guglielmo BJ, Jacobson PA, Kradjan WA, et al. Koda-kimble and Young's applied therapeutics: the clinical use of drugs. 10th ed. Philadelphia, USA: Wolters Kluwer Health Adis (ESP); 2013.
Roberts-Wolfe DJ, Kalivas PW. Glutamate Transporter GLT-1 as a Therapeutic Target for Substance Use Disorders. CNS Neurol Disord Drug Targets. 2015;14(6):745-56. doi: 10.2174/1871527314666150529144655. [PubMed: 26022265]. [PubMed Central: PMC4730383].
Ebrahimie M, Bahmani M, Shirzad H, Rafieian-Kopaei M, Saki K. A Review Study on the Effect of Iranian Herbal Medicines on Opioid Withdrawal Syndrome. J Evid Based Complementary Altern Med. 2015;20(4):302-9. doi: 10.1177/2156587215577896. [PubMed: 25818661].
Motahari AA, Sahraei H, Meftahi GH. Role of Nitric Oxide on Dopamine Release and Morphine-Dependency. Basic Clin Neurosci. 2016;7(4):283-90. doi: 10.15412/J.BCN.03070401. [PubMed: 27872689]. [PubMed Central: PMC5102557].
Listos J, Lupina M, Talarek S, Mazur A, Orzelska-Gorka J, Kotlinska J. The Mechanisms Involved in Morphine Addiction: An Overview. Int J Mol Sci. 2019;20(17). doi: 10.3390/ijms20174302. [PubMed: 31484312]. [PubMed Central: PMC6747116].
Zarrindast M, Mousa-Ahmadi E. Effects of GABAergic system on naloxone-induced jumping in morphine-dependent mice. Eur J Pharmacol. 1999;381(2-3):129-33. doi: 10.1016/s0014-2999(99)00546-4.
Seth V, Upadhyaya P, Moghe V, Ahmad M. Role of calcium in morphine dependence and naloxone-precipitated withdrawal in mice. J Exp Pharmacol. 2011;3:7-12. doi: 10.2147/JEP.S15240. [PubMed: 27186105]. [PubMed Central: PMC4863299].
Shimatani T, Adachi H, Mihashi H, Usumoto N, Yoshimoto K, Ayukawa K. Calcium channel blocker attenuated opioid withdrawal syndrome. Acute Med Surg. 2015;2(2):114-6. doi: 10.1002/ams2.72. [PubMed: 29123703]. [PubMed Central: PMC5667212].
Doyle T, Bryant L, Batinic-Haberle I, Little J, Cuzzocrea S, Masini E, et al. Supraspinal inactivation of mitochondrial superoxide dismutase is a source of peroxynitrite in the development of morphine antinociceptive tolerance. Neuroscience. 2009;164(2):702-10. doi: 10.1016/j.neuroscience.2009.07.019. [PubMed: 19607887]. [PubMed Central: PMC2860430].
Abdel-Zaher AO, Mostafa MG, Farghaly HS, Hamdy MM, Abdel-Hady RH. Role of oxidative stress and inducible nitric oxide synthase in morphine-induced tolerance and dependence in mice. Effect of alpha-lipoic acid. Behav Brain Res. 2013;247:17-26. doi: 10.1016/j.bbr.2013.02.034. [PubMed: 23470902].
Raghavendra V, Tanga FY, DeLeo JA. Attenuation of morphine tolerance, withdrawal-induced hyperalgesia, and associated spinal inflammatory immune responses by propentofylline in rats. Neuropsychopharmacology. 2004;29(2):327-34. doi: 10.1038/sj.npp.1300315. [PubMed: 14532913].
Han Y, Jiang C, Tang J, Wang C, Wu P, Zhang G, et al. Resveratrol reduces morphine tolerance by inhibiting microglial activation via AMPK signalling. Eur J Pain. 2014;18(10):1458-70. doi: 10.1002/ejp.511. [PubMed: 24756886].
Parvardeh S, Moghimi M, Eslami P, Masoudi A. alpha-Terpineol attenuates morphine-induced physical dependence and tolerance in mice: role of nitric oxide. Iran J Basic Med Sci. 2016;19(2):201-8. [PubMed: 27081466]. [PubMed Central: PMC4818369].
Hosseinzadeh H, Parvardeh S, Masoudi A, Moghimi M, Mahboobifard F. Attenuation of morphine tolerance and dependence by thymoquinone in mice. Avicenna J Phytomed. 2016;6(1):55-66. [PubMed: 27247922]. [PubMed Central: PMC4884218].
Sanodiya BS, Thakur GS, Baghel RK, Prasad GB, Bisen PS. Ganoderma lucidum: a potent pharmacological macrofungus. Curr Pharm Biotechnol. 2009;10(8):717-42. doi: 10.2174/138920109789978757. [PubMed: 19939212].
Bishop KS, Kao CH, Xu Y, Glucina MP, Paterson RR, Ferguson LR. From 2000years of Ganoderma lucidum to recent developments in nutraceuticals. Phytochemistry. 2015;114:56-65. doi: 10.1016/j.phytochem.2015.02.015. [PubMed: 25794896].
Kino K, Yamashita A, Yamaoka K, Watanabe J, Tanaka S, Ko K, et al. Isolation and characterization of a new immunomodulatory protein, ling zhi-8 (LZ-8), from Ganoderma lucidium. J Biol Chem. 1989;264(1):472-8. [PubMed: 2909532].
Liu YW, Gao JL, Guan J, Qian ZM, Feng K, Li SP. Evaluation of antiproliferative activities and action mechanisms of extracts from two species of Ganoderma on tumor cell lines. J Agric Food Chem. 2009;57(8):3087-93. doi: 10.1021/jf900011f. [PubMed: 19368349].
Shieh YH, Liu CF, Huang YK, Yang JY, Wu IL, Lin CH, et al. Evaluation of the hepatic and renal-protective effects of Ganoderma lucidum in mice. Am J Chin Med. 2001;29(3-4):501-7. doi: 10.1142/S0192415X01000526. [PubMed: 11789593].
Cui XY, Cui SY, Zhang J, Wang ZJ, Yu B, Sheng ZF, et al. Extract of Ganoderma lucidum prolongs sleep time in rats. J Ethnopharmacol. 2012;139(3):796-800. doi: 10.1016/j.jep.2011.12.020. [PubMed: 22207209].
Sun L, Qiao YL, Sui ZJ, Zhang JY, Song XT, Wu XG. Experimental research on the sleep promoting effects of G. lucidum spore powder. China Practical Med. 2013;8(30):250-1.
Chu QP, Wang LE, Cui XY, Fu HZ, Lin ZB, Lin SQ, et al. Extract of Ganoderma lucidum potentiates pentobarbital-induced sleep via a GABAergic mechanism. Pharmacol Biochem Behav. 2007;86(4):693-8. doi: 10.1016/j.pbb.2007.02.015. [PubMed: 17383716].
Wang SQ, Li XJ, Qiu HB, Jiang ZM, Simon M, Ma XR, et al. Anti-epileptic effect of Ganoderma lucidum polysaccharides by inhibition of intracellular calcium accumulation and stimulation of expression of CaMKII alpha in epileptic hippocampal neurons. PLoS One. 2014;9(7). e102161. doi: 10.1371/journal.pone.0102161. [PubMed: 25010576]. [PubMed Central: PMC4092074].
Cherian E, Sudheesh NP, Janardhanan KK, Patani G. Free-radical scavenging and mitochondrial antioxidant activities of Reishi-Ganoderma lucidum (Curt: Fr) P. Karst and Arogyapacha-Trichopus zeylanicus Gaertn extracts. J Basic Clin Physiol Pharmacol. 2009;20(4):289-308. doi: 10.1515/jbcpp.2009.20.4.289. [PubMed: 20214017].
Lobo V, Patil A, Phatak A, Chandra N. Free radicals, antioxidants and functional foods: Impact on human health. Pharmacogn Rev. 2010;4(8):118-26. doi: 10.4103/0973-7847.70902. [PubMed: 22228951]. [PubMed Central: PMC3249911].
Zhang W, Zhang Q, Deng W, Li Y, Xing G, Shi X, et al. Neuroprotective effect of pretreatment with ganoderma lucidum in cerebral ischemia/reperfusion injury in rat hippocampus. Neural Regen Res. 2014;9(15):1446-52. doi: 10.4103/1673-5374.139461. [PubMed: 25317156]. [PubMed Central: PMC4192946].
Chen X, Veena RK, Ramya H, Janardhanan KK, George V. Gano oil: A novel antinociceptive agent extracted from Ganoderma lucidum inhibits paw oedema and relieves pain by hypnotic and analgesic actions of fatty acid amides. J Ethnopharmacol. 2020;263:113144. doi: 10.1016/j.jep.2020.113144. [PubMed: 32730883].
Sheikholeslami MA, Parvardeh S, Ghafghazi S, Moini Zanjani T, Sabetkasaei M. The Attenuating Effect of Curcumin on Morphine Dependence in Rats: The Involvement of Spinal Microglial Cells and Inflammatory Cytokines. Iran J Pharm Res. 2019;18(Suppl1):198-207. doi: 10.22037/ijpr.2019.111701.13309. [PubMed: 32802100]. [PubMed Central: PMC7393048].
Singh J. Maceration, percolation and infusion techniques for the extraction of medicinal and aromatic plants. Extraction technologies for medicinal and aromatic plants. 67. International Centre for Science and High Technology; 2008. p. 32-5.
Saedi Marghmaleki V, Alaei HA, Azizi Malekabadi H, Pilehvarian A. Effect of Physical Activity on Symptoms of Morphine Addiction in Rats, after and before of Lesion of the mPFC Area. Iran J Basic Med Sci. 2013;16(10):1091-9. [PubMed: 24379967]. [PubMed Central: PMC3874096].
Shou WH, Gong BR, Fang YS, He ZP. Effects of G. lucidum granules on sedation, hypnosis and immune function in mice. Chin Pharm. 2003;12:42-3.
Jiang HT, Ren YH, Yu WY, Zhou QC, Shen B. Study of sleep promoting and immunoregulation activity of G. lucidum extract and combination preparation. Edible Fungi. 2009;3:64-6.
Tejwani GA, Rattan AK, Sribanditmongkol P, Sheu MJ, Zuniga J, McDonald JS. Inhibition of morphine-induced tolerance and dependence by a benzodiazepine receptor agonist midazolam in the rat. Anesth Analg. 1993;76(5):1052-60. doi: 10.1213/00000539-199305000-00025. [PubMed: 8484507].
Tejwani GA, Sheu MJ, Sribanditmongkol P, Satyapriya A. Inhibition of morphine tolerance and dependence by diazepam and its relation to mu-opioid receptors in the rat brain and spinal cord. Brain Res. 1998;797(2):305-12. doi: 10.1016/s0006-8993(98)00416-8. [PubMed: 9666154].
Suzuki T, Tsuda M, Narita M, Funada M, Mizoguchi H, Misawa M. Diazepam pretreatment suppresses morphine withdrawal signs in the mouse. Life Sci. 1996;58(4):349-57. doi: 10.1016/0024-3205(95)02294-5. [PubMed: 8538371].
Sahraei H, Ghoshooni H, Hossein Salimi S, Mohseni Astani A, Shafaghi B, Falahi M, et al. The effects of fruit essential oil of the Pimpinella anisum on acquisition and expression of morphine induced conditioned place preference in mice. J Ethnopharmacol. 2002;80(1):43-7. doi: 10.1016/s0378-8741(02)00012-0. [PubMed: 11891086].
Hosseinzadeh H, Nourbakhsh M. Effect of Rosmarinus officinalis L. aerial parts extract on morphine withdrawal syndrome in mice. Phytother Res. 2003;17(8):938-41. doi: 10.1002/ptr.1311. [PubMed: 13680829].
Hosseinzadeh H, Ziaee T. Effect of Nepeta glomerulosa Boiss. Aerial parts aqueous extract on morphine withdrawal syndrome in mice. Iran J Pharm Sci. 2006;2(1):41-6.
Dogrul A, Zagli U, Tulunay FC. The role of T-type calcium channels in morphine analgesia, development of antinociceptive tolerance and dependence to morphine, and morphine abstinence syndrome. Life Sci. 2002;71(6):725-34. doi: 10.1016/s0024-3205(02)01736-8. [PubMed: 12072160].
Wang SY, Hsu ML, Hsu HC, Tzeng CH, Lee SS, Shiao MS, et al. The anti-tumor effect of Ganoderma lucidum is mediated by cytokines released from activated macrophages and T lymphocytes. Int J Cancer. 1997;70(6):699-705. doi: 10.1002/(sici)1097-0215(19970317)70:6<699::aid-ijc12>3.0.co;2-5. [PubMed: 9096652].
Zhu M, Chang Q, Wong LK, Chong FS, Li RC. Triterpene antioxidants from ganoderma lucidum. Phytother Res. 1999;13(6):529-31. doi: 10.1002/(sici)1099-1573(199909)13:6<529::aid-ptr481>3.0.co;2-x. [PubMed: 10479768].
Song YS, Kim SH, Sa JH, Jin C, Lim CJ, Park EH. Anti-angiogenic and inhibitory activity on inducible nitric oxide production of the mushroom Ganoderma lucidum. J Ethnopharmacol. 2004;90(1):17-20. doi: 10.1016/j.jep.2003.09.006. [PubMed: 14698502].
Lee YH, Kim JH, Song CH, Jang KJ, Kim CH, Kang JS, et al. Ethanol Extract of Ganoderma lucidum Augments Cellular Anti-oxidant Defense through Activation of Nrf2/HO-1. J Pharmacopuncture. 2016;19(1):59-69. doi: 10.3831/KPI.2016.19.008. [PubMed: 27280051]. [PubMed Central: PMC4887753].
Ozek M, Uresin Y, Gungor M. Comparison of the effects of specific and nonspecific inhibition of nitric oxide synthase on morphine analgesia, tolerance and dependence in mice. Life Sci. 2003;72(17):1943-51. doi: 10.1016/s0024-3205(03)00100-0. [PubMed: 12597993].
Abdel-Zaher AO, Abdel-Rahman MS, E. Lwasei FM. Blockade of nitric oxide overproduction and oxidative stress by Nigella sativa oil attenuates morphine-induced tolerance and dependence in mice. Neurochem Res. 2010;35(10):1557-65. doi: 10.1007/s11064-010-0215-2. [PubMed: 20552271].
Zeng XS, Geng WS, Wang ZQ, Jia JJ. Morphine Addiction and Oxidative Stress: The Potential Effects of Thioredoxin-1. Front Pharmacol. 2020;11:82. doi: 10.3389/fphar.2020.00082. [PubMed: 32153403]. [PubMed Central: PMC7047156].
Torkzadeh-Mahani S, Esmaeili-Mahani S, Nasri S, Darvishzadeh F, Naderi R. Ginger Extract Reduces Chronic Morphine-Induced Neuroinflammation and Glial Activation in Nucleus Accumbens of Rats. Addict Health. 2019;11(2):66-72. doi: 10.22122/ahj.v11i2.229. [PubMed: 31321003]. [PubMed Central: PMC6633069].
Zhang H, Largent-Milnes TM, Vanderah TW. Glial neuroimmune signaling in opioid reward. Brain Res Bull. 2020;155:102-11. doi: 10.1016/j.brainresbull.2019.11.012. [PubMed: 31790721]. [PubMed Central: PMC6946383].
Pan Y, Sun X, Jiang L, Hu L, Kong H, Han Y, et al. Metformin reduces morphine tolerance by inhibiting microglial-mediated neuroinflammation. J Neuroinflammation. 2016;13(1):1-12. doi: 10.1186/s12974-016-0754-9. [PubMed: 27855689]. [PubMed Central: PMC5114746].
Guo YJ, Yuan H, Gan SW, Li L. Protective effect of Ganoderma lucidum polysaccharide on the brain tissue of rat model of Alzheimer disease induced by betaamyloid protein fragment (25-35). J Histochem Cytochem. 2006;15:448-51.
Joseph S. Antioxidative and Antiinflammatory Activities of the Chloroform Extract of Ganoderma lucidum Found in South India. Scientia Pharmaceutica. 2009;77(1):111-22. doi: 10.3797/scipharm.0808-17.
Ko HH, Hung CF, Wang JP, Lin CN. Antiinflammatory triterpenoids and steroids from Ganoderma lucidum and G. tsugae. Phytochemistry. 2008;69(1):234-9. doi: 10.1016/j.phytochem.2007.06.008. [PubMed: 17655889].
Tran YH, Nguyen TTT, Nguyen PT, Nguyen KT, Duong CX, Tran HM. Effects of Ganoderma Lucidum Extract on Morphine-Induced Addiction and Memory Impairment in Mice. Biointerface Res Appl Chem. 2021;12(1). doi: 10.33263/BRIAC121.10761084.